Murén E, Rask L
Uppsala Genetic Center, Department of Cell Research, Swedish University of Agricultural Sciences, Sweden.
Planta. 1996;200(4):373-9. doi: 10.1007/BF00231392.
The maturation of the 2S albumin, napin, in Brassica napus L. involves removal of an amino-terminal and an internal propeptide. Pulse-chase experiments with B. napus embryos showed that intermediates are detectable during the pronapin processing. Intact pronapin was expressed by baculovirus in Spodoptera frugiperda insect cells in order to obtain substrate for studying the processing event. Processing of pronapin with a crude B. napus embryo protein extract resulted in several fragments of similar sizes to those of napin heavy and light chains. The character of the major processing activity in the B. napus extract suggested that it was due to an aspartic proteinase. A secondary activity indicated an additional endoproteinase involved in the pronapin processing. Limited proteolysis of pronapin with a purified aspartic proteinase from Hordeum vulgare showed that cleavage occurred exclusively in the prosequences. The cleavage products formed in-vitro requires additional trimming of the propeptides in order to obtain the subunits of mature napin.
甘蓝型油菜中2S清蛋白napin的成熟过程涉及去除氨基末端和内部前肽。对甘蓝型油菜胚胎进行的脉冲追踪实验表明，在原napin加工过程中可检测到中间体。为了获得用于研究加工过程的底物，杆状病毒在草地贪夜蛾昆虫细胞中表达了完整的原napin。用粗制的甘蓝型油菜胚胎蛋白提取物处理原napin，产生了几个大小与napin重链和轻链相似的片段。甘蓝型油菜提取物中主要加工活性的特征表明，这是由于一种天冬氨酸蛋白酶所致。二级活性表明有另一种内肽酶参与原napin的加工。用来自大麦的纯化天冬氨酸蛋白酶对原napin进行有限的蛋白水解表明，切割仅发生在前序列中。体外形成的切割产物需要对前肽进行额外的修剪才能获得成熟napin的亚基。
